title: A High-Temperature, High-Throughput Method for Monitoring Residual Formaldehyde in Vaccine Formulations.
abstract: Formaldehyde has long been used in the chemical inactivation of viral material during vaccine production. Viral inactivation is required so that the vaccine does not infect the patient. Formaldehyde is diluted during the vaccine manufacturing process, but residual quantities of formaldehyde are still present in some current vaccines. Although formaldehyde is considered safe for use in vaccines by the Food and Drug Administration, excessive exposure to this chemical may lead to cancer or other health-related issues. An assay was developed that is capable of detecting levels of residual formaldehyde in influenza vaccine samples. The assay employs incubation of dosage formulation suspensions with hydralazine hydrochloride under mildly acidic conditions and elevated temperatures, where formaldehyde is derivatized to yield fluorescent s-triazolo-[3,4-a]-phthalazine. The assay has been traditionally run by high-performance liquid chromatography, where runtimes of 15 minutes per sample can be expected. Our laboratory has developed a plate-based version that drastically improved the throughput to a runtime of 96 samples per minute. The assay was characterized and validated with respect to reaction temperature, evaporation, stability, and selectivity to monitor residual formaldehyde in various influenza vaccine samples, including in-process samples. Heat transfer and evaporation will be especially considered in this work. Since the assay is plate based, it is automation friendly. The new assay format has attained detection limits of 0.01 µg/mL residual formaldehyde, which is easily able to detect and quantify formaldehyde at levels used in many current vaccine formulations (<5 µg/0.5-mL dose).
authors.affiliation: Department of Food, Bioprocessing and Nutrition Sciences, North Carolina State University, Raleigh, NC, USA.
authors.forename: Kendra D
authors.initials: KD
authors.lastname: Stallings
authors.affiliation: Golden LEAF Biomanufacturing Training and Education Center, North Carolina State University, Raleigh, NC, USA.
authors.forename: Rebecca L
authors.initials: RL
authors.lastname: Kitchener
authors.affiliation: Golden LEAF Biomanufacturing Training and Education Center, North Carolina State University, Raleigh, NC, USA nghentz@ncsu.edu.
authors.forename: Nathaniel G
authors.initials: NG
authors.lastname: Hentz
mesh_terms: D057205:Automation, Laboratory; D002138:Calibration; D002273:Carcinogens / Q000032:analysis* / Q000737:chemistry; D004202:Disinfectants / Q000032:analysis* / Q000737:chemistry; D004339:Drug Compounding; D004340:Drug Contamination / Q000517:prevention & control*; D004355:Drug Stability; D005456:Fluorescent Dyes / Q000032:analysis / Q000737:chemistry; D005557:Formaldehyde / Q000032:analysis* / Q000737:chemistry; D057166:High-Throughput Screening Assays; D006358:Hot Temperature; D006830:Hydralazine / Q000737:chemistry; D006863:Hydrogen-Ion Concentration; D007202:Indicators and Reagents / Q000737:chemistry; D007252:Influenza Vaccines / Q000737:chemistry*; D057230:Limit of Detection; D010793:Phthalazines / Q000032:analysis / Q000737:chemistry; D011786:Quality Control; D015203:Reproducibility of Results; D013050:Spectrometry, Fluorescence; D014230:Triazoles / Q000032:analysis / Q000737:chemistry; D014835:Volatilization
chemical_list: D002273:Carcinogens; D004202:Disinfectants; D005456:Fluorescent Dyes; D007202:Indicators and Reagents; D007252:Influenza Vaccines; D010793:Phthalazines; D014230:Triazoles; C479574:s-triazolo(3,4-a)phthalazine; D005557:Formaldehyde; D006830:Hydralazine
keywords: formaldehyde assay; high-temperature assay; residual formaldehyde; vaccine